Cancer is the second leading cause of death in the United States. Although “cancer” is used to describe many different types of cancer, i.e. breast, prostate, lung, colon, pancreas, each type of cancer differs both at the phenotypic level and the genetic level. The unregulated growth characteristic of cancer occurs when the expression of one or more genes becomes dysregulated due to mutations or epigenetic changes, and cell growth can no longer be controlled.
Growth control genes are often classified in two classes, oncogenes and tumor suppressor genes. Oncogenes are genes whose normal function is to promote cell growth, but only under specific conditions. When an oncogene gains a mutation and loses that specificity control, it promotes growth under a much wider variety of conditions. However, it has been found that for cancer to be truly successful the cancer cell must also acquire mutations in tumor suppressor genes. The normal function of tumor suppressor genes is to stop cellular growth. Examples of tumor suppressors include p53, p16, p21, and APC, all of which, when acting normally, stop a cell from dividing and growing uncontrollably. When a tumor suppressor is mutated or lost, that brake on cellular growth is also lost, allowing cells to now grow without the normal restraints.
EphB3 is a receptor in the ephrin receptor tyrosine kinase family. Presently there are 14 Eph receptors and 9 ephrin ligands known in humans. Ephrin receptors (Ephs) and their ligands, the ephrins, mediate numerous developmental processes, particularly in the nervous system and vascular systems. Ephrins are also known to play a role in tumor development, angiogenesis, metastatic growth and cell survival. Based on their structures and sequence relationships, ephrins are divided into the ephrin-A (EFNA) class, which are anchored to the membrane by a glycosylphosphatidylinositol linkage, and the ephrin-B (EFNB) class, which are transmembrane proteins. The Eph family of receptors is divided into 2 groups based on the similarity of their extracellular domain sequences and their affinities for binding ephrin-A and ephrin-B ligands. Eph receptors make up the largest subgroup of the receptor tyrosine kinase (RTK) family.
Eph receptors have been implicated in cancer. NIH3T3 cells transfected with EphA1 and transplanted into nude mice produce 10 mm3 tumors in 5-6 weeks, while vector controls did not produce any tumors during the same time period (Maru et al., Oncogene. 1990 March; 5(3):445-7). EphB2 is expressed at higher levels in cancers of the stomach (12/16), colon (3/11), esophagus (3/6), ovarian (1/7), kidney (1/2) and lung (1/1) when compared to normal tissues (Kiyokawa et al., Cancer Res 1994 Jul. 15; 54(14):3645-50). EphB6 expression correlates with low grade neuroblastomas. The kinase domain of EphB6 is not active, therefore this receptor has been proposed to act as a naturally occurring dominant negative (Tang et al., Clin Cancer Res 1999a Jun; 5(6):1491-6).
There is an increasing volume of evidence that implicates the involvement of Ephs and ephrins in angiogenesis. EphrinA1, ephrinB1, ephrinB2, EphB2, EphB3 and EphB4 have been reported to be expressed in blood vessels. EphrinA1 can induce angiogenesis in the rat cornea model and antibodies to ephrinA1 can inhibit TNF-α induced angiogenesis in this same model (Pandey et al., Science 1995 Apr. 28; 268(5210):567-9). Clustered ephrinB1 induces cell attachment and capillary-like assembly in P19, a teratocarcinoma-derived murine cell line, and in human renal microvascular endothelial cells (HRMEC) (Stein et al., Genes Dev 1998 Mar. 1; 12(5):667-78). Clustered ephrinB1 and ephrinB2 can also induce sprouting of adrenal-cortex derived microvascular endothelial cells (ACE) (Adams et al., Genes Dev 1999 Feb. 1; 13(3):295-306). Injection of a dominant negative EphB4 receptor RNA in Xenopus embryos causes intersomitic veins to project abnormally into adjacent somites (Helbling et al., Development 2000 January; 127(2):269-78). EphB2/EphB3 double knockout, EphB4 and ephrinB2 knockout mice all have vascular remodeling defects (Adams et al., 1999; Wang et al., Cell 1998 May 29; 93(5):741-53; PCT WO 00/30673).
Ephs may also play a role in metastasis. 293T human epithelial kidney cells transfected with either EphB3 or EphB2 exhibit reduced cell adhesion to fibronectin or collagen coated surfaces in vitro. Failure of 293 cells to adhere was mediated by EphB2 phosphorylation of R-ras followed by integrin de-activation (Zou et al., Proc Natl Acad Sci USA 1999 Nov. 23; 96(24):13813-8).
Ephs appear to function by signaling upon activation. Ephrin binding induces Eph receptor oligomerization causing phosphorylation of juxtamembrane residues of Ephs. Activated Ephs have multiple phosphorylated tyrosines that act as docking sites for signaling proteins (e.g. RasGAP, Src, LMW-PTP, PLCg, PI3-kinase, Grb2, and PDZ containing proteins).
Overexpression of Eph receptors (EphA1, EphA2, EphB2) causes transformation in the absence of receptor hyper-phosphorylation. Phosphorylated EphB receptors negatively regulate the Ras-MAP-kinase pathway and FAK signaling, impairing cell growth.
EphB3 (also known as Hek2, Sek4, Mdk5, Tyro6, Cek10 and Qek10) is a receptor for ephrin B family members (ephrin B1, ephrin B2 and ephrin B3), and is known to be expressed in normal tissue and in certain tumors and cancer cell lines. To date, however, the role of EphB3 in cancer has not been elucidated. Thus, there is a need to identify compositions and methods that modulate EphB3 and its role in such cancers. The present invention is directed to these, as well as other, important needs.